Curacin A is a mixed polyketide/non-ribosomal peptide with antimitotic properties produced by the marine cyanobacterium Lyngbya majuscula (4). The hybrid polyketide synthase (PKS)/non-ribosomal peptide synthase (NRPS) biosynthetic pathways that produces curacin A (5) contains numerous unique chemical steps, many of which have been previously investigated (1,3,6,22). The synthesis of a terminal alkene, instead of the carboxyl typical for this class of linear natural products, and the unique domain arrangement in the terminal PKS module are mysteries yet to be fully elucidated.
In CurM, the terminal module, a sulfotransferase (ST) and thioesterase (TE) domain follow the acyl carrier protein (ACP) (FIG. 1a.) STs catalyze the transfer of a sulfate group from the donor 3′-adenosine 5′-phosphosulfate (PAPS) to a hydroxyl or amine of their acceptor small molecule or protein (10). STs are involved in vital processes such as detoxification, hormone regulation, and signaling and have been studied in a wide variety of organisms. However, the ST in the curacin pathway is the first observation of a ST within a PKS module and contains low (15%) sequence identity to other studied STs.
The TE, although identifiable as a thioesterase, does not resemble any of the previously established fatty acid synthase (FAS), PKS or NRPS TE families (24). Many PKS offloading TEs have been studied to date including the TEs of the pikromycin synthase (Pik) (25,26) and erythromycin synthase (DEBS) (27) pathways. PKS offloading TEs typically perform either hydrolysis to produce a carboxylic acid or catalyze the attack of an intramolecular hydroxyl to form a macrolactone. These TEs are dimers, with two N-terminal alpha helices forming a lid-to-lid dimer interface, and adopt the α/β hydrolase fold characteristic of some serine hydrolases. Access to the classic nucleophile-His-acid catalytic triad active site is restricted by a narrow tunnel formed by a closed lid. Many PKS and NRPS pathways also include a second non-modular thioesterase called a TE II (in addition to an offloading TE, also known as TE I), which performs an editing function within the pathway. The curacin TE shows low similarity to sequences in all parts of the phylogenic tree (24), also pointing the need to more closely study curacin TE to understand its activity.